Struelens M J, De Gheldre Y, Deplano A
Université Libre de Bruxelles, Belgium.
Infect Control Hosp Epidemiol. 1998 Aug;19(8):565-9. doi: 10.1086/647874.
A number of high-resolution molecular typing systems have been developed in recent years. Their availability raises the new issues of selecting the method (s) best suited for a particular purpose and interpreting and communicating typing results. Most of the currently available methods are comparative only: they allow testing of a sample of isolates for delineation of those closely related from those markedly different in genomic backgrounds. This approach is adequate for outbreak investigation, allowing determination of clonal spread in a microenvironment and identification of the source of infection. Comparative methods with sufficient resolution for most pathogens include restriction fragment-length polymorphism (RFLP), pulsed-field gel electrophoresis (PFGE), and arbitrarily primed and randomly amplified polymorphic DNA-polymerase chain reaction (PCR) analysis. For surveillance systems, monitoring clonal spread and prevalence in populations over extended periods of time requires library typing systems. These must be standardized, must have a high throughput, and must use a uniform nomenclature. Promising or validated methods include serotyping, insertion sequence fingerprinting, ribotyping, PFGE, amplified fragment-length polymorphism (AFLP), infrequent-restriction-site amplification PCR, interrepetitive element PCR typing (rep-PCR) and PCR-RFLP of polymorphic loci. PCR methods generating arrays of size-specific amplicons (AFLP, rep-PCR) can be more reproducibly analyzed by using denaturing polyacrylamide gel or capillary electrophoresis with automated laser detection. Binary probe typing systems appear optimal and should be enhanced further through use of DNA chip technology. In these systems, amplification of polymorphic regions is followed by solid-phase hybridization with a reference panel of sequence-variant specific probes. The resulting binary type results allow determination of reproducible, numeric profiles. However, interpretation and nomenclature of typing results for large-scale surveillance purposes still require a better understanding of population structure and microevolution of most microbial pathogens.
近年来已开发出多种高分辨率分子分型系统。这些系统的出现引发了一些新问题，即如何选择最适合特定目的的方法以及如何解读和交流分型结果。目前大多数可用方法仅具有比较性：它们允许对分离株样本进行检测，以区分基因组背景密切相关的菌株和明显不同的菌株。这种方法适用于暴发调查，可确定微环境中的克隆传播并识别感染源。对大多数病原体具有足够分辨率的比较方法包括限制性片段长度多态性（RFLP）、脉冲场凝胶电泳（PFGE）以及任意引物随机扩增多态性DNA聚合酶链反应（PCR）分析。对于监测系统，在较长时间内监测人群中的克隆传播和流行情况需要文库分型系统。这些系统必须标准化，必须具有高通量，并且必须使用统一的命名法。有前景或经过验证的方法包括血清分型、插入序列指纹图谱分析、核糖体分型、PFGE、扩增片段长度多态性（AFLP）、低频限制性位点扩增PCR、重复元件间PCR分型（rep-PCR）以及多态性位点的PCR-RFLP。通过使用变性聚丙烯酰胺凝胶或具有自动激光检测功能的毛细管电泳，可以更可重复地分析产生大小特异性扩增子阵列的PCR方法（AFLP、rep-PCR）。二元探针分型系统似乎是最佳的，应通过使用DNA芯片技术进一步加强。在这些系统中，多态性区域的扩增之后是与序列变异特异性探针的参考面板进行固相杂交。由此产生的二元分型结果可用于确定可重复的数字图谱。然而，对于大规模监测目的的分型结果的解读和命名仍需要更好地了解大多数微生物病原体的群体结构和微观进化。
